Project Summary ? Project 1 Elucidating biomarkers and mechanisms of the Ketogenic longevity mechanism. The isocaloric Ketogenic Diet (KD) initiated in middle-aged mice reprograms metabolism, decreases p46Shc and preserves late-life functions including memory, cognition, and many parameters of muscle function, and significantly increases lifespan1,2. A better understanding of the ketosis-dependent longevity mechanism and its biomarkers could lead to more effective dosing of this nutritional intervention for its delay of aging and cognitive decline in mice, and by extension in humans. Our goals include identifying the KD's longevity mechanism and biomarkers, by RNAseq and protein expression in tissues and blood (Aims1 and 2). By identifying the genes which increase or decrease the most, we identify the most KD-responsive biomarkers. By overlapping these biomarkers with the results of Proj 2(Pelicci), which will determine the aging-relevant epigenetic marks and which aging-relevant epigenetic marks are reversed by KD, we will identify those biomarkers that are both aging-relevant and KD-responsive. These will help to mechanistically define genes that support the KD's preservation of functional longevity. These aging-relevant and KD-responsive biomarkers will also serve Proj4 Ramsey's goals to evaluate the relative benefit of iKD and KS strategies of 'therapeutic ketosis with respect to functional longevity'. In Aim 3, we test three likely candidate mechanisms for KD's longevity effect, i.e. rising ketones that target the beta-hydroxybutyrate receptor HCAR2, FFAR3, and the repression of p46Shc that regulates thiolase and ketogenesis. Reversal of functional longevity phenotype will indicate prominent involvement of HCAR2, FFAR3 and p46Shc in the mechanism, which can be further specified into by the mitochondrial and acetylation and KAT mechanisms by Project 3 (Baar). In Aim 4, we test recently-identified Shc inhibitors in a cell model of Alzheimer's, and the PSAPP mouse model of Alzheimer's disease (With Core C). The completion of this integrative set of experiments will define aging-relevant biomarkers of the KD, and will test three likely mechanisms of the KD explicitly. It will also determine whether small-molecule Shc inhibitors protect in cell and animal models of AD, which could lead to a new pharmacological strategy for Alzheimer's disease not based on dissolution of amyloid, but based on a neuro-metabolic, mitochondrial and pro-survival strategy. The overall product of the Project will support a mechanistic understanding of what therapeutic ketosis is with respect to functional longevity, how it works, what its biomarkers are, and potential benefit of Shc inhibitors in AD.